Apoptosis, or programmed cell death, plays a central role in the development and homeostasis of all multicellular organisms. A frequent hallmark of cancer is resistance to natural apoptotic signals. Depending on the cancer type, this resistance can be due to up- or down-regulation of key proteins in the apoptotic cascade. The resistance can also be due to mutations in genes encoding these proteins. These changes can occur in both the intrinsic apoptotic pathway, which funnels through the mitochondria and caspase-9, and the extrinsic apoptotic pathway, which involves the action of death receptors and caspase-8. For example, alterations in healthy levels of proteins such as p53, Bim, Bax, Apaf-1, FLIP and many others have been observed in cancers. These alterations can lead to a defective apoptotic cascade, one in which the upstream proapoptotic signal is not adequately transmitted to activate the executioner caspases, caspase-3 and caspase-7.
As most apoptotic pathways ultimately involve the activation of procaspase-3, upstream genetic abnormalities are effectively “breaks” in the apoptotic circuitry, and as a result such cells proliferate atypically. Given the central role of apoptosis in cancer, efforts have been made to develop therapeutics that target specific proteins in the apoptotic cascade. However, because these therapeutics target early (or intermediate to high) positions on the apoptotic cascade, cancers with mutations in proteins downstream of those members can still be resistant to the possible beneficial effects of those compounds.
For therapeutic purposes, it would be advantageous to identify small molecules that directly activate a proapoptotic protein far downstream in the apoptotic cascade. This approach could involve a relatively low position in the cascade, thus enabling the killing of even those cells that have mutations in their upstream apoptotic machinery. Moreover, such therapeutic strategies would have a higher likelihood of success if that proapoptotic protein were upregulated in cancer cells. Thus, identifying small molecules that target the downstream effector protein of apoptosis, procaspase-3, would significantly aid current cancer therapy.
The conversion or activation of procaspase-3 to caspase-3 results in generation of the active “executioner” caspase form that subsequently catalyzes the hydrolysis of a multitude of protein substrates. In certain cancers, the levels of procaspase-3 are elevated relative to normal tissue. A study of primary isolates from 20 colon cancer patients revealed that on average, procaspase-3 was upregulated six-fold in such isolates relative to adjacent non-cancerous tissue. In addition, procaspase-3 is upregulated in certain neuroblastomas, lymphomas, and liver cancers. Furthermore, a systematic evaluation of procaspase-3 levels in the 60 cell-line panel used for cancer screening by the National Cancer Institute Developmental Therapeutics Program was performed. The evaluation revealed that certain lung, melanoma, renal, and breast cancers show greatly enhanced levels of procaspase-3 expression. Due to the role of active caspase-3 in achieving apoptosis, the relatively high levels of procaspase-3 in certain cancerous cell types, and the intriguing safety catch-mediated suppression of its autoactivation, small molecules that directly modify procaspase-3, could have great applicability in targeted cancer therapy.
Furthermore, combination therapy has become increasingly important for the treatment of cancer patients. The goal of combination therapy is to achieve an additive or synergistic effect between chemotherapeutics, thereby facilitating shortened treatment times, decreased toxicity, and increased patient survival. Drugs that act on a single biochemical pathway are particularly strong candidates for synergy or potentiation as they may mimic “synthetic lethal” genetic combinations. Thus, there is an urgent a need for more effective therapies for the treatment of many forms of cancer, and new synergistic combinations of anticancer drugs would aid this pursuit. Accordingly, there exists a need to identify new combinations of cytotoxic agents that are effective in killing cancer cells, yet protect normal host tissues from the undesired toxicity of the cytotoxic agent.